In the application of biological tissu e engineering,three-dimensional and longitudinal evaluation of biological struc tural markers is needed.However,these biological structures are usually several mi llimeters thick and cloudy,thus,it is very challenging for imaging,and classical fluorescence microscopy can not meet its needs. Therefore,we develo ped a mesoscopic fluorescence molecular tomography method.Mesoscopic fluorescenc e molecular imaging system was a new imaging system,which filled the gap between microfluoresce nce molecular imaging technology and macrofluorescence molecular imaging technology. In order to improve the performance of mesoscopic fluorescence molecular reconstruct ion,this paper mainly improved the configuration of the system.The configuration paramete rs of the optical system,including detector layout,uncoupled or coupled scanning mode,were optimized based on optical principles,and the 3d imaging performance of the mesos copic fluorescent molecular imaging system was evaluated and compared.The result s show that the coupling backlight mesoscopic fluorescence molecular tomography imaging (MFMT) system designed in this paper can improve the reconstruction performance and obtain high-qualit y reconstruction results.